T cell proliferation in response to mitogens or activation of the T cell receptor represents a crucial component of the immune response. It is well established that this proliferative response declines as a function of age in rodents, as well as humans, but the mechanisms involved are as yet poorly understood. We have previously shown that hepatic expression of thiostatin, a strong inhibitor of cysteine proteinases, greatly increases during senescence of rats in vivo. This may affect the rate of cell division, since inhibition of cysteine proteinases is known to disrupt cell cycle progression. Our preliminary data suggest that thiostatin overexpression in vitro (in a mouse fibroblast model) indeed interferes with both cell division and signal transduction. While the liver is essentially post-mitotic, other groups have shown that serum thiostatin does get internalized by a variety of cells. Preliminary data from our laboratory indicates that thiostatin does indeed accumulate intracellularly in splenocytes from old rats. This accumulation is paralleled by changes in the activity and steady state levels of proteins involved in MAPK dependent signal transduction which are reminiscent of the effects observed in fibroblasts upon overexpression of thiostatin. We hypothesize that inhibition of cysteine proteinases by thiostatin during aging results in an imbalance in the levels of key proteins within the cell, and that this imbalance in turn impairs the cell~s ability to respond to mitogenic stimuli. Furthermore, we hypothesize that inhibition of proteolysis by thiostatin might account, at least partially, for the decreased proliferative ability of T lymphocytes obtained from old individuals. Therefore, we propose to directly test this hypothesis by examining the ability of thiostatin to reduce the Con A-induced proliferative response of young lymphocytes. The aged (>65) are the fastest growing segment of our population, and the process of immunosenescence is believed to have significant implications for the health and well-being of older citizens. We believe this is an appropriate time to address some of the molecular defects involved in this phenomenon, since both the technology and the basic understanding of the activity of the relevant pathways are currently in place.